A Primer to Angiotensin Peptide Isolation, Stability, and Analysis by Nano-Liquid Chromatography with Mass Detection

Methods in Molecular Biology
Mariola OlkowiczRyszard T Smolenski

Abstract

The renin-angiotensin system (RAS) is an important element of cardiovascular and renal physiology and targeting the RAS by renin inhibitors, angiotensin (Ang) converting enzyme (ACE) inhibitors and Ang II type 1 receptor antagonists is effective in the treatment of hypertension, heart failure, and atherosclerosis. Quantification of Ang peptides is critical to establish the status of the RAS, but it is challenging due to low Ang peptides concentrations (fmol/mL or fmol/g), abundance of interfering substances, post sampling conversions, and difficulties with the specificity of the assay.In this chapter, we describe a new nano-LC/MS-based methodology for comprehensive, specific, sensitive, and accurate quantification of Ang peptides profile in plasma and tissue. We optimized sample pretreatment method (protein removal (acetonitrile precipitation) followed by solid-phase extraction (C18 silica bonded phase)), chromatographic conditions (reversed-phase nanochromatography with preconcentration), and mass detection (multiple reaction monitoring) of nine peptides: Ang-(1-12), Ang I (1-10), Ang-(1-9), Ang II (1-8), [Ala1]-Ang II, Ang III (2-8), Ang IV (3-8), Ang-(1-7), and [Ala1]-Ang-(1-7). Assessment of plasma and cardiac concentration...Continue Reading

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